Rheumatic Disease Sera: Probes of Disease Mechanism

描述(由申请人提供)：这项提案的广泛、长期目标是定义人类自身免疫性肌病的致病机制，以期确定这类疾病的治疗途径。最近的几项研究已经确认细胞毒性淋巴细胞颗粒通路在自身免疫性肌炎中具有潜在的重要作用。在多发性肌炎中，细胞毒性淋巴细胞颗粒经常偏向肌肉细胞，表明这些淋巴细胞处于脱颗粒的活跃过程中。此外，针对多发性肌炎和皮肌炎的所有自身抗原都统一于它们对颗粒酶B(GRB)有效切割的敏感性，产生在其他形式的细胞死亡过程中不产生的独特片段。我们假设，细胞毒性淋巴细胞颗粒途径在诱导肌肉细胞死亡和产生独特形式的自身抗原方面具有双重作用，这些自身抗原驱动自身免疫性肌炎的自身免疫反应。该提案的具体目的是(1)确定细胞毒性淋巴细胞在体外诱导肌肉细胞功能障碍和死亡的机制。这将通过在体外阐明GRB下游的效应通路，并通过确定在成肌细胞中观察到的显著抑制caspase的机制来实现；(2)确定自身免疫性肌炎患者体内受影响组织中的肌肉细胞损伤的途径。目标1中被证明具有功能相关性的GRB下游通路的活性将使用一套专门报告GRB、半胱氨酸酶、钙蛋白酶、nNOS以及肌肉结构和调节蛋白的状态和活性的新型试剂来直接询问；(3)在动物模型中阐明GRB介导的切割和肌肉细胞分化状态对肌肉抗原免疫原性的作用。这些研究将探讨GRB的切割在产生对肌炎特异性自身抗原EF-1α的特异性免疫反应中的作用，并确定成肌细胞是否是肌肉特异性自身抗原的自身抗体反应的优先启动者。综上所述，这些研究将加深我们对自身免疫性肌炎发病机制的理解。

DESCRIPTION (provided by applicant): The broad, long-term objectives of this proposal are to define pathogenic mechanisms in human autoimmune myopathies, with a view to defining pathways of therapeutic relevance in this group of diseases. Several recent studies have identified the cytotoxic lymphocyte granule pathway as a pathway of potential importance in autoimmune myositis. In polymyositis, cytotoxic lymphocyte granules are frequently polarized towards muscle cells, indicating that these lymphocytes are in the active process of degranulation. Furthermore, all autoantigens targeted in polymyositis and dermatomyositis are unified by their susceptibility to efficient cleavage by granzyme B (GrB), generating unique fragments not generated during other forms of cell death. We hypothesize that the cytotoxic lymphocyte granule pathway plays a dual role in inducing muscle cell death and in generating the unique forms of autoantigens, which drive the autoimmune response in autoimmune myositis. The specific aims of the proposal are to (1) Define the mechanisms whereby cytotoxic lymphocytes induce muscle cell dysfunction and death in muscle cells in vitro. This will be accomplished by elucidating the effector pathways downstream of GrB in muscle cells in vitro, and by defining the mechanisms responsible for the prominent inhibition of caspases observed in myoblasts; (2) Define the pathways of muscle cell damage in vivo in affected tissues from patients with autoimmune myositis. The activity of the pathways downstream of GrB demonstrated to be of functional relevance in Aim 1 will be directly interrogated using a set of novel reagents that specifically report on the state and activity of GrB, caspases, calpains, nNOS, and muscle structural and regulatory proteins; (3) Elucidate the role of GrB-mediated cleavage and muscle cell differentiation state on the immunogenicity of muscle antigens in an animal model. These studies will address the role of cleavage by GrB in generating the specific immune response to the myositis-specific autoantigen EF-1alpha and define whether the myoblast is the preferential initiator of autoantibody responses to muscle-specific autoantigens. Taken together, these studies will enhance our understanding of the effector mechanisms that participate in the pathogenesis of autoimmune myositis.